title: Innocuous full-length botulinum neurotoxin targets and promotes the expression of lentiviral vectors in central and autonomic neurons.
abstract: Fragments of botulinum neurotoxin (BoNT) have been explored as potential targeting moieties and carriers of biomolecules into neurons, although with lower binding and translocation efficiency compared with intact proteins. This study exploits a detoxified recombinant form of full-length BoNT/B (BoTIM/B) fused with core streptavidin (CS-BoTIM/B) for lentiviral targeting to central and autonomic neurons. CS-BoTIM/B underwent an activity-dependent entry into cultured spinal cord neurons. Coupling CS-BoTIM/B to biotinylated lentivirus-encoding green fluorescent protein (GFP) endowed considerable neuron selectivity to the vector as evident from the preferential expression of the reporter in neurons co-cultured with skeletal muscle cells. CS-BoTIM/B-guided lentiviral transduction with the expression of a SNARE protein, SNAP-25 (S25), rendered non-susceptible to proteolysis by three BoNT serotypes, yielded a sizable decrease in cleaved S25 upon exposure of spinal cord neurons to these toxins. This was accompanied by synaptic transmission being spared from blockade by BoNT/A or BoNT/E, reflecting adequate translation and functional competence of recombinant multi-toxin-resistant S25. The augmented neurotropism conveyed on the lentivirus by CS-BoTIM/B was also demonstrated in vivo through enhanced expression of a reporter in intramural ganglionic neurons in the rat trachea, after injection of the targeted GFP-encoding lentivirus. Thus, a novel and realistic prospect for gene therapy of peripheral neuropathies is offered in this study through lentiviral targeting to neurons by CS-BoTIM/B.
authors.affiliation: International Centre for Neurotherapeutics, Dublin City University, Dublin, Ireland.
authors.forename: V B
authors.initials: VB
authors.lastname: O'Leary
authors.forename: S V
authors.initials: SV
authors.lastname: Ovsepian
authors.forename: A
authors.initials: A
authors.lastname: Raghunath
authors.forename: Q
authors.initials: Q
authors.lastname: Huo
authors.forename: G W
authors.initials: GW
authors.lastname: Lawrence
authors.forename: L
authors.initials: L
authors.lastname: Smith
authors.forename: J O
authors.initials: JO
authors.lastname: Dolly
mesh_terms: D001905:Botulinum Toxins / Q000494:pharmacology*; D019274:Botulinum Toxins, Type A; D005725:Ganglia, Autonomic / Q000378:metabolism*; D018390:Gene Targeting; D018014:Gene Transfer Techniques; D005822:Genetic Vectors; D049452:Green Fluorescent Proteins / Q000235:genetics; D007395:Interneurons / Q000378:metabolism*; D016086:Lentivirus / Q000235:genetics*; D009928:Organ Specificity; D011994:Recombinant Proteins / Q000494:pharmacology; D013116:Spinal Cord / Q000166:cytology / Q000378:metabolism*; D019809:Streptavidin; D009435:Synaptic Transmission; D050825:Synaptosomal-Associated Protein 25 / Q000378:metabolism
chemical_list: D011994:Recombinant Proteins; D050825:Synaptosomal-Associated Protein 25; C096323:rimabotulinumtoxinB; D049452:Green Fluorescent Proteins; D019809:Streptavidin; D001905:Botulinum Toxins; D019274:Botulinum Toxins, Type A